S-Homocysteinylated Proteins - Homocysteine in Protein Structure/Function and Human Disease - page 124

7.2.2 Fibronectin

The assembly of fibrillin-1 depends on fibronectin, which exists in two forms:
soluble plasma fibronectin, synthesized by hepatocytes, and cellular fibrous form
secreted and assembled by mesenchymal cells [435]. Cellular fibronectin is
secreted as a disulfide-bonded dimer with

60 cysteine residues, most of which
form intrachain disulfides. In addition to fibrillin, fibronectin also assembles other
matrix proteins: collagen types I and III, thrombospondin-1, fibulin-1, and LTBP-1.
In vitro experiments show that when human plasma is treated with 0.5-mM [ 35 S]
Hcy, plasma fibronectin, in addition to albumin, undergoes S-homocysteinylation.
Purified fibronectin also binds [ 35 S]Hcy, and the binding is reversed by the treat-
ment with the reducing agent 2-mercaptoethanol [436]. About five Hcy molecules
bind per each 440-kDa fibronectin dimer. Mass spectroscopic analyses indicate that
S-homocysteinylated regions are located in the N- and C-terminal domains, but not
in the collagen-binding region [436].
However, the treatment with 0.5 mM [ 35 S]Cys does not lead to fibronectin S-
cysteinylation (in plasma or purified), which indicates that the binding is specific
for Hcy. The incorporation of Hcy via disulfide linkages affects fibronectin func-
tion, as shown by the inhibition of S-Hcy-fibronectin binding to fibrin. However,
the binding to gelatin/collagen, involving a different domain of fibrillin, is not
affected by S-homocysteinylation [436].
Fibronectin S-homocysteinylation compromises its interaction with fibrillin-1
but not with heparin [437]. Hcy, but not cysteine, reduces the fibronectin disulfide-
bound dimer to monomers and modifies epitopes for disulfide-dependent binding of
monoclonal antifibronectin antibodies.
S-homocysteinylation inhibits de novo assembly of fibronectin on cells [437].
For example, the treatment with 1-mM Hcy reduces, while 5-mM Hcy completely
abolishes, fibronectin deposition on human dermal fibroblasts. Similar treatments
with cysteine do not affect fibronectin assembly while the treatment with DTT
completely prevented fibronectin deposition and network formation on fibroblasts.
These results indicate that native disulfide bonds in fibronectin molecule are
important for its function and that the disruption of these disulfide bonds impairs
normal fibronectin function. It remains to be determined, however, whether
S-homocysteinylation of fibronectin or other components of the extracellular matrix
occurs in CBS-deficient patients or in human hyperhomocysteinemia in general.

>

7.2.3 Tropoelastin

Microfibrils formed by fibrillins serve as a scaffold for the deposition of
tropoelastin, which is important for the formation and maintenance of elastic fibers.
Properly assembled elastic fibers confer elasticity on tissues such as skin, lung,
and aorta. During elastic fiber biogenesis, tropoelastin undergoes maturation and